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Gene expression profiling reveals similarities between the spatial architectures of postnatal articular and growth plate cartilage.

ABSTRACT: Articular and growth plate cartilage are discrete tissues but arise from a common cartilaginous condensation and have comparable spatial architectures consisting of distinct layers of chondrocytes. To investigate similarities and differences between articular and growth plate cartilage and to explore transcriptional changes that occur during the onset of their divergence, we performed manual microdissection of 10-day-old rat proximal tibias, microarray analysis, bioinformatics, and real-time PCR to compare gene expression profiles in individual cartilage layers. We found that many genes that were spatially upregulated in the intermediate/deep zone of articular cartilage were also spatially upregulated in the resting zone of growth plate cartilage (overlap greater than expected by chance, P<0.001). Interestingly, the superficial zone of articular cartilage showed an expression profile with similarities to both the proliferative and hypertrophic zones of growth plate cartilage (P<0.001 each). Additionally, significant numbers of known proliferative zone markers (3 out of 6) and hypertrophic zone markers (27 out of 126) were spatially upregulated in the superficial zone (more than expected by chance, P<0.001 each). In conclusion, we provide evidence that the intermediate/deep zone of articular cartilage has a gene expression profile more similar to that of the resting zone of growth plate cartilage, whereas the superficial zone has a gene expression profile more similar to those of the proliferative and hypertrophic zones. These findings suggest that the superficial zone chondrocytes of articular cartilage differentiate according to a program that is not completely different from but instead has distinct similarities to the hypertrophic differentiation program of growth plate chondrocytes. We also present functional signaling pathways implicated by differential gene expression between articular and growth plate cartilage during their initial separation by the secondary ossification center.

SUBMITTER: Chau M

PROVIDER: S-EPMC4113381 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Gene expression profiling reveals similarities between the spatial architectures of postnatal articular and growth plate cartilage.

Chau Michael M Lui Julian C JC Landman Ellie B M EB Späth Stephan-Stanislaw SS Vortkamp Andrea A Baron Jeffrey J Nilsson Ola O

PloS one 20140728 7

Articular and growth plate cartilage are discrete tissues but arise from a common cartilaginous condensation and have comparable spatial architectures consisting of distinct layers of chondrocytes. To investigate similarities and differences between articular and growth plate cartilage and to explore transcriptional changes that occur during the onset of their divergence, we performed manual microdissection of 10-day-old rat proximal tibias, microarray analysis, bioinformatics, and real-time PCR ...[more]

PMID: 25068449

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Project description:Articular and growth plate cartilage both arise from condensations of mesenchymal cells, but ultimately develop important histological and functional differences. Each is composed of three layers-the superficial, mid and deep zones of articular cartilage and the resting, proliferative and hypertrophic zones of growth plate cartilage. The bone morphogenetic protein (BMP) system plays an important role in cartilage development. A gradient in expression of BMP-related genes has been observed across growth plate cartilage, likely playing a role in zonal differentiation. To investigate the presence of a similar expression gradient in articular cartilage, we used laser capture microdissection (LCM) to separate murine growth plate and articular cartilage from the proximal tibia into their six constituent zones, and used a solution hybridization assay with color-coded probes (nCounter) to quantify mRNAs for 30 different BMP-related genes in each zone. In situ hybridization and immunohistochemistry were then used to confirm spatial expression patterns. Expression gradients for Bmp2 and 6 were observed across growth plate cartilage with highest expression in hypertrophic zone. However, intracellular BMP signaling, assessed by phospho-Smad1/5/8 immunohistochemical staining, appeared to be higher in the proliferative zone and prehypertrophic area than in hypertrophic zone, possibly due to high expression of Smad7, an inhibitory Smad, in the hypertrophic zone. We also found BMP expression gradients across the articular cartilage with BMP agonists primarily expressed in the superficial zone and BMP functional antagonists primarily expressed in the deep zone. Phospho-Smad1/5/8 immunohistochemical staining showed a similar gradient. In combination with previous evidence that BMPs regulate chondrocyte proliferation and differentiation, the current findings suggest that BMP signaling gradients exist across both growth plate and articular cartilage and that these gradients may contribute to the spatial differentiation of chondrocytes in the postnatal endochondral skeleton.

Project description:Metaphyseal chondrodysplasia, Schmid type (MCDS) is characterized by mild short stature and growth plate hypertrophic zone expansion, and caused by collagen X mutations. We recently demonstrated the central importance of ER stress in the pathology of MCDS by recapitulating the disease phenotype by expressing misfolding forms of collagen X (Schmid) or thyroglobulin (Cog) in the hypertrophic zone. Here we characterize the Schmid and Cog ER stress signaling networks by transcriptional profiling of microdissected mutant and wildtype hypertrophic zones. Both models displayed similar unfolded protein responses (UPRs), involving activation of canonical ER stress sensors and upregulation of their downstream targets, including molecular chaperones, foldases, and ER-associated degradation machinery. Also upregulated were the emerging UPR regulators Wfs1 and Syvn1, recently identified UPR components including Armet and Creld2, and genes not previously implicated in ER stress such as Steap1 and Fgf21. Despite upregulation of the Chop/Cebpb pathway, apoptosis was not increased in mutant hypertrophic zones. Ultrastructural analysis of mutant growth plates revealed ER stress and disrupted chondrocyte maturation throughout mutant hypertrophic zones. This disruption was defined by profiling the expression of wildtype growth plate zone gene signatures in the mutant hypertrophic zones. Hypertrophic zone gene upregulation and proliferative zone gene downregulation were both inhibited in Schmid hypertrophic zones, resulting in the persistence of a proliferative chondrocyte-like expression profile in ER-stressed Schmid chondrocytes. Our findings provide a transcriptional map of two chondrocyte UPR gene networks in vivo, and define the consequences of UPR activation for the adaptation, differentiation, and survival of chondrocytes experiencing ER stress during hypertrophy. Thus they provide important insights into ER stress signaling and its impact on cartilage pathophysiology.

Project description:BackgroundArticular cartilage (AC) is the layer of tissue that covers the articulating ends of the bones in diarthrodial joints. Across species, adult AC shows an arcade-like structure with collagen predominantly perpendicular to the subchondral bone near the bone, and collagen predominantly parallel to the articular surface near the articular surface. Recent studies into collagen fibre orientation in stillborn and juvenile animals showed that this structure is absent at birth. Since the collagen structure is an important factor for AC mechanics, the absence of the adult Benninghoff structure has implications for perinatal AC mechanobiology. The current objective is to quantify the dynamics of collagen network development in a model animal from birth to maturity. We further aim to show the presence or absence of zonal differentiation at birth, and to assess differences in collagen network development between different anatomical sites of a single joint surface. We use quantitative polarised light microscopy to investigate properties of the collagen network and we use the sheep (Ovis aries) as our model animal.ResultsPredominant collagen orientation is parallel to the articular surface throughout the tissue depth for perinatal cartilage. This remodels to the Benninghoff structure before the sheep reach sexual maturity. Remodelling of predominant collagen orientation starts at a depth just below the future transitional zone. Tissue retardance shows a minimum near the articular surface at all ages, which indicates the presence of zonal differentiation at all ages. The absolute position of this minimum does change between birth and maturity. Between different anatomical sites, we find differences in the dynamics of collagen remodelling, but no differences in adult collagen structure.ConclusionsThe collagen network in articular cartilage remodels between birth and sexual maturity from a network with predominant orientation parallel to the articular surface to a Benninghoff network. The retardance minimum near, but not at, the articular surface at all ages shows that a zonal differentiation is already present in the perinatal animals. In these animals, the zonal differentiation can not be correlated to the collagen network orientation. We find no difference in adult collagen structure in the nearly congruent metacarpophalangeal joint, but we do find differences in the dynamics of collagen network remodelling.

Project description:BackgroundArticular cartilage undergoes an important maturation process from neonate to adult that is reflected by alterations in matrix protein organization and increased heterogeneity of chondrocyte morphology. In the horse, these changes are influenced by exercise during the first five months of postnatal life. Transcriptional profiling was used to evaluate changes in articular chondrocyte gene expression during postnatal growth and development.MethodsTotal RNA was isolated from the articular cartilage of neonatal (0-10 days) and adult (4-5 years) horses, subjected to one round of linear RNA amplification, and then applied to a 9,367-element equine-specific cDNA microarray. Comparisons were made with a dye-swap experimental design. Microarray results for selected genes (COL2A1, COMP, P4HA1, TGFB1, TGFBR3, TNC) were validated by quantitative polymerase chain reaction (qPCR).ResultsFifty-six probe sets, which represent 45 gene products, were up-regulated (p < 0.01) in chondrocytes of neonatal articular cartilage relative to chondrocytes of adult articular cartilage. Conversely, 586 probe sets, which represent 499 gene products, were up-regulated (p < 0.01) in chondrocytes of adult articular cartilage relative to chondrocytes of neonatal articular cartilage. Collagens, matrix-modifying enzymes, and provisional matrix non-collagenous proteins were expressed at higher levels in the articular cartilage of newborn foals. Those genes with increased mRNA abundance in adult chondrocytes included leucine-rich small proteoglycans, matrix assembly, and cartilage maintenance proteins.ConclusionDifferential expression of genes encoding matrix proteins and matrix-modifying enzymes between neonates and adults reflect a cellular maturation process in articular chondrocytes. Up-regulated transcripts in neonatal cartilage are consistent with growth and expansion of the articular surface. Expression patterns in mature articular cartilage indicate a transition from growth to homeostasis, and tissue function related to withstanding shear and weight-bearing stresses.

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Gene expression profiling reveals similarities between the spatial architectures of postnatal articular and growth plate cartilage.

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Gene expression profiling reveals similarities between the spatial architectures of postnatal articular and growth plate cartilage.

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